Development of a Universal Influenza Vaccine Against Influenza A and B Viruses Influenza virus causes widespread mortality and morbidity every year. In addition, the threat of an influenza pandemic continues to persist. For current seasonal licensed vaccines to be effective the influenza strain in the formulation should match that which is circulating in the human population. Unfortunately, making a prediction of the influenza strains that are likely to circulate in the human population in the future is not very reliable. As a result, an error in this prediction can make the vaccine ineffective. This unreliability of the vaccine exists because the vaccine is based on one of the most abundant membrane proteins called hemagglutinin found on the influenza virus surface. Because hemagglutinin changes from one strain to the next, a proper match between circulating influenza strains and that in the vaccine is important. Furthermore, because the identity of a future pandemic strain cannot be predicted, it is hard to develop a vaccine for pandemic influenza based on hemagglutinin's head region as an antigen. To overcome the limitation of the current vaccine design we propose to use highly conserved antigens to formulate an influenza vaccine. One of these conserved antigens is from the ion channel membrane protein called matrix 2 (M2). The domain of M2 exposed on the surface of the virus is called M2e, and it has remained highly conserved in human influenza A strains. By attaching consensus human M2e on the gold nanoparticle surface we have shown breadth of protection against H1N1 and H3N2 influenza A strains, and even the highly pathogenic avian influenza strain H5N1. The vaccine was however only partially protective against the highly pathogenic avian influenza A H7N9 strain. The reason is that M2e on avian and swine influenza strains shows some dissimilarity from M2e of human influenza strains. Therefore, we propose that inclusion of M2e of human, avian and swine influenza strains as the vaccine antigen will increase the breadth of protection. The second conserved antigen is an epitope from the neuraminidase membrane protein of the influenza virus. This epitope is conserved across influenza A and B strains. We hypothesize that inclusion of both M2e and the conserved neuraminidase epitope will help to design a universal influenza vaccine protective against a broad range of strains. Our specific aims are: AIM 1: Develop the multi-antigen vaccine formulation, and establish its breadth and longevity of protection in Balb/c mice. AIM 2: Characterize the role of humoral and cellular immunity in the mechanism of protection, and assess biodistribution and safety profile of the vaccine. AIM 3: Establish vaccine efficacy in ferrets, and evaluate vaccine thermal stability. The influenza vaccine designed through this research is expected to have a broad and significant impact on public health. If successful, the vaccine will offer broad protection against both influenza A and B strains, eliminating the need for seasonal vaccines, and significantly reducing the threat of pandemic outbreaks due to influenza virus.